Guided tissue regeneration (GTR) is a surgical procedure which uses Gore- tex membranes or Gore-tex augmentation membranes (GTAM) to favor the repopulation of the healing wound with cells which have bone regenerative potential. Regenerative tissue is a unique environment in which the process of pathological resorption is converted to bone regeneration. Inhibition of osteoclast-mediated resorption may be as critical to regeneration as the stimulation of osteoblast-mediated bone synthesis, due to the processes being tightly coupled. The activity of both synthetic and degradative cells is controlled by factors such as cytokines. Our preliminary results have demonstrated that conditioned medium from cells derived from regenerative tissue inhibits osteoclast differentiation. The specific aims of this project are (1) to determine the magnitude of the inhibition of osteoclast differentiation and resorptive activity induced by cells from different regenerative tissues; (2) to isolate the factor(s) from the regenerative cells which mediates inhibition; and (3) to determine whether the factor(s) has (have) identity with known cytokines (interferon- gamma, interleukin-4, transforming growth factor-beta), which inhibit osteoclast differentiation and resorptive activity. The focus will be on the cell lines established from tissue adherent to Gore-tex membranes, removed from patients following GTR and cells adherent to GTAM membranes, recovered from edentulous ridge augmentation procedures. Conditioned medium from the regenerative cell lines will be tested for activity in the osteoclast-like cell differentiation and the osteoclast resorption assays. Both assays are necessary as one assesses the number of osteoclasts formed from hemopoietic precursors and the other measures the amount of bone resorbed by these cells. The conditioned medium from the regenerative cell lines will be fractionated and partially purified, and the fractions tested for inhibitory activity in the bioassays. To determine whether these factors are known cytokines, fractions will be probed with antibodies to specific cytokines on Western blots and ELISA. Whether the cytokines are the cause of the inhibition will be determined by conducting bioassays with blocking antibodies to the cytokines and by testing whether this factor can synergize with known cytokines to produce the inhibitory effect.